Probe Summary for Inhibitors and Stabilizers of Firefly Luciferase

Firefly (Photinus pyralis) luciferase is one of the most commonly used transcriptional reporters. It is considered one of the more dynamically responsive reporters - that is, responsive to changes in reporter transcription - as it requires no post-translational modifications and is enzymatically active directly after protein synthesis (Wood, 1998). With no endogenous expression in mammalian cells and without the requirement of external excitation, it offers very little background signal, thus generally providing excellent signal to background for most assays. ..more

Firefly (Photinus pyralis) luciferase is one of the most commonly used transcriptional reporters. It is considered one of the more dynamically responsive reporters - that is, responsive to changes in reporter transcription - as it requires no post-translational modifications and is enzymatically active directly after protein synthesis (Wood, 1998). With no endogenous expression in mammalian cells and without the requirement of external excitation, it offers very little background signal, thus generally providing excellent signal to background for most assays.

Previous profiling work to determine the prevalence of compounds that affect FLuc enzymatic activity (PubChem AID 411) identified that ~3% of the MLSMR library (then at ~72K) inhibited FLuc (Auld et al., 2008). More recent profiling efforts using KM concentrations of FLuc substrates indicate that the number of compounds that inhibit FLuc is even greater than this estimate. In cell-based FLuc reporter gene assays, it has been found that compounds that inhibit FLuc can cause either inhibition or activation of FLuc (Thorne et al., 2010). Whether inhibition or activation of FLuc is demonstrated in a cell-based assay by a FLuc inhibitory compound depends on the potency of the compound, the mode of inhibition of the compound, the assay format and protocol, and the detection reagent used. FLuc inhibitory compounds that cause activation in a cell-based FLuc reporter gene assay have been found to interact with the FLuc protein in the cell, thereby stabilizing it, increasing its protein half-life, and allowing it to accumulate intracellularly (Thompson et al., 1991). An example of such compounds are a subset of 3,5-diaryl oxiadiazoles that are potent FLuc inhibitors that can increase the melting temperature of FLuc (Auld et al., 2010).